Expression of the ras oncogene p21 product by hepatocytes of cirrhotic liver with hepatocellular carcinoma (HCC) was examined immunohistochemically using mouse monoclonal antibody RAP-5. At the concentration of antibody used, histologically normal liver tissues were negative for p21 antigen, whereas hepatocytes of cirrhotic nodules from 80 of 92 HCC patients (87.4%), and 10 of 32 patients without HCC (59.4%) were positive. This difference was statistically significant (p less than 0.05). The incidence of p21 expression by hepatocytes was significantly higher in macronodular cirrhotic patients than in those with micronodular cirrhosis (p less than 0.05) and tended to be higher in those positive for hepatic hepatitis B virus markers than in those that were negative (p less than 0.1). All 16 patients with liver cell dysplasia, and 17 of 18 with adenomatous hyperplasias showed increased expression of p21 antigen. In HCC it was detected on tumor cells of 63 of 101 patients (62.4%). Characteristically, its expression in well-differentiated HCC was mild and uniformly diffuse, and in moderately differentiated tumors was markedly heterogeneous in both intensity and distribution, whereas no expression was observed in cells of poorly differentiated HCC. These observations suggest that elevated ras p21 antigen expression is important in the development of both cirrhotic nodules and HCC, but that after tumor development, its sustained elevation is no longer necessary for cell proliferation and progression through the grades of anaplasia.